Nicotine is the agent responsible for dependency to tobacco. But how does nicotine dependency, and the often-severe pathological consequences, develop in some, but not all, users? Nicotine exerts its effect(s) through specific ion-channel receptors termed nicotinic acetylcholine receptors (nAChRs). Because these receptors are the biological entry point through which nicotine exert its effects, understanding their expression and role in modulating cellular functions is imperative. However, the subunit composition and expression of native nAChRs is poorly defined as are the cellular mechanisms through which nicotine pre-conditions and modifies cellular responses. In this project we address these issues experimentally using genetically defined mouse strains with varying susceptibility to nicotine to define the distribution of key nAChR subunits in the brain. We will use microarray technology to define the impact of nicotine on transcription activation by the inflammatory cytokine, TNF alpha, and define proteolytic signaling cascades and responses that are altered by nicotine. Finally, we implement state-of-the-art gene-targeting and replacement methods to test directly the participation of key genes and/or pathways that are hypothesized to contribute to the genetic predisposition to nicotine as outlined below. Specific Aim 1. Strain-specific nAChR subunit distribution and expression. Hypothesis 1: The relative expression of nAChR subunit protein differs between inbred mouse strains. Hypothesis 2: Chronic nicotine (oral or smoking) alters region-specific nAChR subunit expression differently between inbred mouse strains. Specific Aim 2. Nicotine alters inflammatory transcriptional responses. Hypothesis: Strain-specific responses to nicotine pre-conditioning alter transcriptional response(s) induced by the pro-inflammatory cytokine, tumor necrosis alpha (TNFa). Specific Aim 3. Nicotine alters inflammatory post-transcriptional responses. Hypothesis: Strain specific responses to nicotine pre-conditioning modify protease activation and/or function induced by the proinflammatory cytokine, tumor necrosis alpha (TNF(x). Specific Aim 4. Strain specific zene-replacement of candidate genes. Hypothesis: Allelic replacement, through homologous recombination, of nAChR candidate gene polymorphisms from the C3H mouse into the C57BU6 will confer a novel phenotype on the recipient as defined in Specific Aims 1-3.